Methods for electrical or electrochemical detection of molecular interactions between biomolecules have provided an attractive alternative to detection techniques relying on radioactive or fluorescent labels. Electrical and electrochemical detection techniques are based on the detection of alterations in the electrical properties of an electrode arising from interactions between molecules attached to the surface of the electrode (often referred to as “probe” molecules) and molecules present in a reaction mixture (often referred to as “target” molecules) in contact with the electrode. Examples of methods and devices related to electrical or electrochemical detection of biomolecules are disclosed in U.S. Pat. Nos. 4,072,576, 4,098,645, 4,414,323, 4,840,893, 5,164,319, 5,187,096, and 5,891,630.
Electrical or electrochemical detection eliminates many of the disadvantages inherent in the use of radioactive or fluorescent labels to detect interactions between probe and target molecules. Electrical or electrochemical detection is safe, inexpensive, sensitive, and is not burdened with complex and onerous regulatory requirements. The development of microfabricated arrays (microarrays) of bio- and chemical molecules has led to further improvements of traditional analytical techniques. Microarrays of bio- and molecules (e.g. oligonucleotides, nucleic acids, proteins, peptides, or antibodies) have utility in a wide variety of applications in which molecular interactions between target molecules in a reaction mixture and large numbers of distinct probe molecules bound to defined regions of a substrate can be simultaneously assayed using electrical, optical, or radioactive detection strategies. Microarrays address the demands for inexpensive, high-throughput detection of biomolecular interactions. Obviously, microarrays can also provide a low cost and high-throughput platform for detection of chemical species. There are, however, problems associated with electrical or electronic systems as such devices cannot detect the same way as optical scanners do and require multiplexing to scan different detection spots. Electronic detection systems also require an electrolyte (either solid or liquid), making ionic isolation of different detection spots critical.
Although biochip arrays for the electrochemical detection of molecular interactions between bio- and/or chemical molecules have been proposed, these devices have significant disadvantages. For example, the device disclosed by Egger et al. in U.S. Pat. Nos. 5,670,322 and 5,532,128 cannot be made column-and-row (or “x-y”) addressable, thus limiting the density of the test sites in the array and the usefulness of the apparatus. In U.S. Pat. No. 5,653,939, Hollis et al. disclosed an x-y addressable array wherein a solid supporting substrate comprises a plurality of test sites in electrochemical contact with a set of orthogonally oriented electrodes. However, Hollis et al. does not provide an apparatus for eliminating the ionic shortage when an electrolyte is applied in microarray chips, in which case, the array electrodes cannot be addressed.
US Patent Application No. 20020090649A1 discloses a high density addressable array for electronic detection biochips. The array, however, does not provide for true four-terminal electronic detection for high S/N ratio and high sensitivity, and does not lend itself for use in high density array chips without isolation channels.
Impedance measurements could be used to directly detect the impedance changes after biomolecular interaction. Most protein and DNA sensors require labelling (such as florescent labels) to report biomolecular interactions. Impedance can enable label-less detection. However, the sensitivity needs to be further improved.
Electronic or electrochemical sensors require two electrodes (working and counter/reference electrodes) or three electrodes (working, counter and reference electrodes) for detection measurements. Existing electronic array sensors are designed to have either two or three electrodes at every array detection site, or are designed to have common counter or reference electrodes. Two or three electrodes per test site typically results in many electrodes and high manufacturing cost. Common counter or reference electrodes cause different solution resistance resulting in additional noise and difficult data processing.
There thus remains a need for an improved column-and-row addressable chem/bio chip array for the electrical or electrochemical detection of molecular interactions that are effective while reducing the cost of performing various analyses, and that can also be easily and cost effectively fabricated. Such devices, and methods for their use, may have wide application in the fields of medicine and genetics and molecular biology research.